Stapling of side chains to stabilize an α-helical structure has been generally associated with an increased uptake of CPPs. Here, we compare four amphiphilic stapled peptides with their linear counterparts in terms of their membrane binding and conformational features in order to correlate these with uptake efficiency and toxicological effects. The impact of lactam stapling was found to vary strongly with regard to the different aspects of peptide-membrane interactions. Nearly all stapled peptides caused less membrane perturbation (vesicle leakage, hemolysis, bacterial lysis) than their linear counterparts. In one case (MAP-1) where stapling enhanced α-helicity in aqueous and lipid environments, leakage was eliminated while cell uptake in HEK293 and HeLa cells remained high, which improved the overall characteristics. The other systems (DRIM, WWSP, KFGF) did not improve, however. The data suggest that cell uptake of amphipathic CPPs correlates with their adopted α-helix content in membranes rather than their helicity in solution.

Five series (28 structures) of photoswitchable β-hairpin peptides were synthesized based on the cyclic scaffold of the natural antibiotic gramicidin S. Cell-type selectivity was compared for all activated (diarylethene "ringopen") and deactivated ("ring-closed") forms in terms of antibacterial activity (MIC against Escherichia coli and Bacillus subtilis), anticancer activity (IC 50 against HeLa cell line), and hemolytic cytotoxicity (HC 50 against human erythrocytes). Correlations between the conformational plasticity of the peptides, their hydrophobicity, and their bioactivity were also analyzed. Considerable improvements in selectivity were achieved compared to the reference compound. We found a dissociation of the anticancer activity from hemolysis. Phototherapeutic indices (PTI), HC 50 (closed)/MIC(open) and HC 50 (closed)/IC 50 (open), were introduced for the peptides as safety criteria. The highest PTI for HeLa-selective toxicity were observed among analogues containing hydroxyleucine on the hydrophobic face. For one compound, high PTIs were demonstrated across a range of different cancer cell lines, including a doxorubicin-resistant one.

Diarylethene derivatives, the biological activity of which can be reversibly changed by irradiation with light of different wavelengths, have shown promise as scientific tools and as candidates for photocontrollable drugs. However, examples demonstrating efficient photocontrol of their biological activity are still relatively rare. This concept article discusses the possible reasons for this situation and presents a critical analysis of existing data and hypotheses in this field, in order to extract the design principles enabling the construction of efficient photocontrollable diarylethene-based molecules. Papers addressing biologically relevant interactions between diarylethenes and biomolecules are analyzed; however, in most published cases, the efficiency of photocontrol in living systems remains to be demonstrated. We hope that this article will encourage further discussion of design principles, primarily among pharmacologists, synthetic and medicinal chemists.

We report the photophysical properties of diarylethene‐based photoswitches (DAE) which were incorporated into the backbones of macrocyclic peptides of variable sizes (6, 10 and 14 amino acids: cDAE‐6, cDAE‐10, and cDAE‐14). The insertion leads to a consistent bathochromic shift in UV/Vis absorption bands, a considerable peptide‐size‐dependent fluorescence intensity increase (up to a factor of 1.7), and a drastically reduced photoisomerization efficiency leading to formation of the ring‐closed DAE fragment. The compounds were studied by time‐resolved photoinduced broadband absorption spectroscopy, revealing a ring closure reaction for the unconstrained DAE reference system and, to a lesser extent, for cyclic cDAE‐14. The smallest cyclic peptide cDAE‐6 does not undergo any photoisomerization process at all. Furthermore, the DAE fragment ring closure reaction in the reference system is concurrent with intersystem crossing that is complete within a few picoseconds. Further analysis showed dominant triplet absorption throughout the observed spectral window (350–700 nm) in all macrocyclic systems. As a consequence, ring closure is favored in the unconstrained system in contrast to the cyclic peptides. Our studies demonstrate the importance of understanding the population dynamics of DAE rotamers when designing efficiently photocontrollable diarylethene‐based peptidomimetics.

A bicyclic peptide scaffold was chemically adapted to generate diarylethene‐based photoswitchable inhibitors of serine protease Bos taurus trypsin 1 (T1). Starting from a prototype molecule—sunflower trypsin inhibitor‐1 (SFTI‐1)—we obtained light‐controllable inhibitors of T1 with Ki in the low nanomolar range, whose activity could be modulated over 20‐fold by irradiation. The inhibitory potency as well as resistance to proteolytic degradation were systematically studied on a series of 17 SFTI‐1 analogues. The hydrogen bond network that stabilizes the structure of inhibitors and possibly the enzyme–inhibitor binding dynamics were affected by isomerization of the photoswitch. The feasibility of manipulating enzyme activity in time and space was demonstrated by controlled digestion of gelatin‐based hydrogel and an antimicrobial peptide BP100‐RW. Finally, our design principles of diarylethene photoswitches are shown to apply also for the development of other serine protease inhibitors.